This report includes work arising from the following clinical protocols: NCT00026832, NCT00100360, NCT00001177, and NCT00001322. Previous studies have identified the presence of abnormalities in resting state activity and reward processing in mood disorders. Indeed, altered resting state function represents an intriguing possible explanation for the generation of ovarian hormone-related state changes characterized by predictable alterations in social, cognitive (e.g., autobiographical memory), and affective function in women with PMDD. Results from our resting state H215O PET studies in healthy premenopausal women (n = 40) demonstrate effects of both hypogonadism and ovarian steroids on resting rCBF in several brain regions, including BA25, insula, and medial PFC (mPFC). Evidence also suggests that genomic variation in sex-steroid receptors or in the systems regulated by sex steroids may contribute to the ovarian hormone-sensitivity observed in PMDD. Thus, as a first step, we examined the effects of common genetic variations in women during each of the hormonal conditions established within the GnRH agonist-induced hypogonadism and ovarian steroid addback protocol. We have tested specific, common functional variants (BDNF Val66Met and COMT Val158Met) that are regulated by sex steroids and that are known to alter brain function. In our studies, therefore, we employ these gene variants as probes for the impact of genetic substrates on the effects of ovarian steroids on womens brain function. We have previously identified that variations in the BDNF gene, which is modulated by ovarian hormones and contains an estrogen response element (ERE), differentially affect working memory-related functions only in the presence of estradiol and only in the hippocampus in healthy women (Wei et al. 2018). Similarly, the gene coding for Catechol-O-Methyltransferase (COMT) contains both EREs and progesterone response elements (PREs) in its promoter region, its expression in brain can be regulated by estradiol (and in some tissues by progesterone) and COMT activity is increased by circulating estradiol in women. Indeed, two studies in women reported the modulatory effects on working memory-related brain regions of both estradiol and COMT genotype (Dumas et al. 2018, Jacobs and D'Esposito 2011). To further understand the interactions between ovarian steroids and BDNF/COMT genotypes on brain circuits related to hippocampal and PFC activity, we used oxygen-15 water regional cerebral blood flow (rCBF) positron emission tomography (PET) to study healthy women who participated the GnRH agonist hormone manipulation study. Preliminary findings demonstrate an ovarian hormone-by-genotype interaction in the hubs of the working memory network with BDNF genotype and estradiol regulating activity in hippocampus, and COMT genotype and estradiol regulating activity within the PFC. These data suggest that the effects of sex hormones on working memory-related brain function are mediated by genotypic variations in a brain region-specific manner. In a companion collaboration focusing on the BDNF gene with the Rockefeller University, we examine the effects of ovariectomy and estradiol replacement on both behavior and hippocampal gene expression in the female humanized BDNF heterozygous Met mouse. We observed that heterozygous Met mice show a differential behavioral sensitivity to estradiol in several hippocampal tasks and in anxiety-like behaviors. Additionally, whole transcriptome RNA sequencing in the BDNF heterozygous Met mouse (hippocampus) and in women with PMDD (lymphoblastoid cell lines) show a significant overlap in the expression of several gene networks stimulated by estradiol. No similar overlap in gene expression was observed after estradiol exposure in either wild type mice or asymptomatic control women. This cross-species overlap in gene expression suggests the potential physiologic (and behavioral) relevance of the BDNF heterozygote met mouse to our understanding of PMDD. Overall, the findings from these studies demonstrate that in healthy women, variations in genes relevant to ovarian steroids can impact functional neurocircuitry and network level gene expression in regionally- and hormonally-specific ways. Specifically, interactions between BDNF and COMT genotypes and estradiol on the activation of functional neurocircuitry in the hippocampus (where BDNF is highly expressed) and the DLPFC (where COMT has primacy for dopamine trafficking), respectively. In studies currently underway, we will examine these gene-hormone interactions in reward-responsive and resting state neural circuits. Our functional genomic studies in cell lines from both women with and without PMDD have demonstrated the relevance of our prior finding of abnormal function of the ESC/E(Z) gene complex in PMDD as follows: First, neuroprogenitor cells (NPCs) were successfully differentiated from induced pluripotent stem cells (iPSCs) made from women with PMDD and control women (confirmed by immunofluorescent staining and transcriptome analysis). Overlapping gene expression between lymphoblastoid cells (LCLs) and NPCs reveals that several genes and gene pathways potentially important for PMDD pathophysiology are expressed in both. Indeed, both LCLs and NSCs show signatures of differential gene silencing through similar mechanisms at baseline and in response to ovarian steroids. These findings in NSCs reveal neuronal differences between women with PMDD and controls both at baseline and after ovarian steroids, including ESC/E(Z) targets. Second, we employed two stock neuronal cell lines: Luhmes Cells, which are immortalized neuronal precursor cell lines (both undifferentiated/neuronal precursors and differentiated/mature dopaminergic neurons), as well as SH-SY5Y, an immortalized neuroblastoid cell line characterized by an adrenergic and dopaminergic phenotype. Both stock neuronal cell lines contained evidence of ESC/E(Z) gene complex function and responsivity to ovarian steroids. Thus, these data provide strong supportive evidence of the neural relevance of our initial findings in lymphoblastoid cell lines (immune-origin tissue) in several more pathophysiologically-relevant tissues (i.e., human NPCs, two stock neuronal cell lines).